Our long-term goals are to devise vectors for efficient gene transfer in cells and intact animals. We have exploited the unique properties of retroviruses to generate high efficiency vectors capable of delivering foreign genetic material upon infection of recipient cells. Specifically, we plan to undertake the following experiments for the coming year: 1) Generation of high efficiency vectors. Attempts will be made to generate retroviruses containing heterologous promoters. We currently have vectors containing HPRT, DHFR, ADA, PNP, factor IX and growth hormone genes. 2) Tissue-specific enhancers. We propose to introduce tissue specific enhancers in the retroviral vectors. 3) Helper-free cell lines. Cell lines capable of packaging infectious virus particles containing foreign genes will be generated. 4) Bone marrow infections. We plan to introduce foreign genes into mouse bone marrow cells and transplant them into mice. Attempts will also be made to establish in vitro long-term human bone marrow cultures. We would also infect a variety of human hematopoietic cell lines. 5) Infection of fibroblasts. As an alternative approach to infection of somatic cells, we plan to infect mouse fibroblasts and transplant them under the skin. A genetically deficient mouse (lit/lit) will be used as a model system. 6) Infection of embryonal cells. Attempts will be made to infect EC cells with retroviral vectors containing two genes, one linked to LTR and another transcribed to an heterologous promoter. 7) Germline transfer. We plan to generate a number of transgenic mice by infecting preimplantation embryos with recombinant retroviruses. Attempts will also be made to introduce proto-oncogenes in the germ line. 8) Expression libraries. We plan to use retroviral vectors to generate expression cDNA libraries. The resultant retroviruses will be used to infect a wide variety of cell types. The long-term ability of retroviral vectors is their potential use in human somatic cell gene therapy. We hope the experiments proposed here will form a strong base for such an approach.